In this paper, to further demonstrate the importance of surface electrostatic interactions to the kinetic structural stabilities of proteins, we perform studies on one hyperthermophile-derived triosephosphate isomerase (TIM) and a homologous psychrophile-derived TIM, transplanting the residues of what appears to be a key helix in the protein, between the two homologs, forming six ion-pairs in the hyperthermophile homolog and only two ion-pairs in the psychrophile homolog. We show that by performing such engineering of proteins, one can modulate the kinetic structural thermal stability of the entire protein, to either lower it, or raise it. The implications of this demonstration are profound, because such electrostatic interaction engineering can be performed to make proteins more thermally stable, or less stable, through changes in kinetic stability rather than thermodynamic stability.

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